Molecular extensibility of mini-dystrophins and a dystrophin rod construct

被引:44
作者
Bhasin, N
Law, R
Liao, G
Safer, D
Ellmer, J
Discher, BM
Sweeney, HL
Discher, DE [1 ]
机构
[1] Univ Penn, Penn Muscle Inst, Philadelphia, PA 19104 USA
[2] Univ Penn, Grad Grp Phys, Philadelphia, PA 19104 USA
[3] Univ Penn, Grad Grp Cell & Mol Biol, Philadelphia, PA 19104 USA
关键词
dystrophin; spectrin; AFM; protein folding; hinge;
D O I
10.1016/j.jmb.2005.07.064
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Muscular dystrophies arise with various mutations in dystrophin, implicating this protein in force transmission in normal muscle. With 24 three-helix, spectrin repeats interspersed with proline-rich hinges, dystrophin's large size is an impediment to gene therapy, prompting the construction of mini-dystrophins. Results thus far in dystrophic mice suggest that at least one hinge between repeats is necessary though not sufficient for palliative effect. One such mini-dystrophin is studied here in forced extension at the single molecule level. Delta 2331 consists of repeats (R) and hinges (H) H1-R1-2 similar to H3 similar to R22-24-H4 linked by native. (-) and non-native (similar to) sequence. This is compared to its core fragment R2 similar to H3 similar to R22 as well as an eight-repeat rod fragment middle (RFM: R8-15). We show by atomic force microscopy that all repeats extend and unfold at forces comparable to those that a few myosin molecules can generate. The hinge regions most often extend and transmit force while limiting tandem repeat unfolding. From 23-42 degrees C, the dystrophin constructs also appear less temperature-sensitive in unfolding compared to a well-studied I-spectrin construct. The results thus reveal new modes of dystrophin flexibility that may prove central to functions of both dystrophin and mini-dystrophins. (c) 2005 Elsevier Ltd. All rights reserved.
引用
收藏
页码:795 / 806
页数:12
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